Immunotherapy has shown promise for the treatment of malignancy. Both cytolytic T cells and lymphokines can facilitate tumor cell destruction, and strategies to enhance tumor regression by administration of cytokines or activated lymphocytes have shown efficacy in animal models and human trials. Recently, we have developed a novel approach to the immunotherapy of malignancy through modification of tumor cells by direct gene transfer in vivo. This approach differs from previous methods, in which tumor cells are propagated, modified, and selected in vitro and subsequently introduced back into the host. In a mouse model, we have used either retroviral vectors or DNA/liposome complexes to deliver recombinant genes to tumor cells in vivo. A gene encoding a highly immunogenic molecule, an allogeneic class I major histocompatibility complex (MHC) glycoprotein, has been expressed in transplantable mouse tumors. When a foreign MHC gene is expressed on these tumors in vivo, it induces specific cytolytic T cells against this protein and also elicits a response against parental unmodified tumor cells, causing tumor regression or cure in animals. This approach has been recently approved for a human phase I trial at the University of Michigan. In this proposal, the introduction of a foreign histocompatibility gene directly within tumors to elicit their immunotherapeutic destruction will be optimized and the efficacy of DNA liposome complexes compared to viral vectors. The expression of allogeneic MHC will be assessed in transduced human melanomas, as well as the immunologic and antitumor effects. This analysis will also be used to develop systems which will allow molecular cloning of immunologically relevant tumor associated antigens in the future. Finally, we will develop modifications of the current protocol which will improve the efficacy of this form of treatment in man. To optimize efficacy, we will; 1) develop methods of pre-immunization to enhance the immune response to in vivo gene transfer, 2) utilize exogenous cytokines to modulate the immune response to gene modified tumors, 3) perform adoptive T cell transfer to further enhance the antitumor response, 4) examine methods to improve selectivity of gene transfer by catheter delivery systems and antibody targeted DNA liposome complexes, and 5) explore the use of other genes alone or in combination with HLA-B7. These studies will allow the development of new modifications of our current human clinical trial which will improve its efficacy and provide insight into the molecular basis of immune recognition in malignancy.